1. Field of the Invention
The present invention relates to cancer therapy, angiogenesis inhibitors, and methods of use thereof. In particular, the present invention describes vascular endothelial growth factor (VEGF) receptor antagonists and their ability to inhibit angiogenesis.
2. Background of the Invention
Vascular endothelial growth factor (VEGF) is a well characterized pro-angiogenic factor (Larrivee, et al., Intl. J. Mol. Med., 5: 447-456 (2000)). VEGF was purified initially from conditioned media of folliculostellate cells and from a variety of tumor cell lines (Ferrara and Henzel, Biochem. Biophys. Res. Comm., 161:851-858 (1989); Myoken et al., Proc Natl Acad Sci USA., 88:5819-23(1991)). VEGF is a member of the cysteine-knot family of growth factors, which also includes PDGF (Platelet Derived Growth Factor). Recently, a number of VEGF structural homologs have been identified: VEGF-B, VEGF-C, VEGF-D and Placenta Growth Factor (PlGF) (Klagsbrun and D'Amore, Cytokine Growth Factor Rev., 7:259-70 (1996); reviewed in Ferrara et al., Endocr. Rev. 18:4-25 (1997).
The human gene encoding VEGF is organized into eight exons, separated by seven introns. Alternative splicing of mRNAs for the VEGF gene results in the generation of five different molecular species, having 121, 145, 165, 189, or 206 amino acid residues in the mature monomer (Tisher et al., 1991; Houck et al., Mol Endocrinol., 5:1806-14 (1991). Only VEGF165, which lacks the residues encoded by exon 6, is the mature and active form of VEGF. It binds to heparin and cell surface heparan sulfate proteoglycans, and can be expressed as a free or a cell membrane bound form (Houck et al., J Biol. Chem., 267:26031-7 (1992).
The endothelial proliferative activity of VEGF is known to be mediated by two high affinity tyrosine kinase receptors, flt-1 (VEGFR1) and KDR (VEGFR2), which exist only on the surface of vascular endothelial cells (DeVries, et al., Science, 225:989-991 (1992) and Terman, et al., Oncogene 6:1677-1683 (1991)). Both the flt-1 and KDR tyrosine kinase receptors have seven immunoglobulin-like (Ig1-7) domains which form the extracellular ligand-binding regions of the receptors, a transmembrane domain which serves to anchor the receptor on the surface of cells in which it is expressed and an intracellular catalytic tyrosine kinase domain which is interrupted by a “kinase insert.” While the KDR receptor binds only the VEGF protein with high affinity, the flt-1 receptor also binds placenta growth factor. An additional member of the receptor tyrosine kinases having seven Ig-like domains in the extracellular ligand-binding region is flt-4, which is not a receptor for either VEGF or PlGF, but instead binds to a different ligand: VH1.4.5. The VH1.4.5 ligand has been reported in the literature as VEGF-related protein (VRP) or VEGF-C (Lee et al., PNAS U.S.A., 93:1988-92 (1996); Ferrara et al., (1997)).
The molecular mechanism by which VEGF initiates signaling is thought to be through receptor dimerization followed by receptor autophosphorylation (see, Ferrara et al., Nat. Med. 9:669-676 (2003)). In other words, VEGF exerts its biological effects on responsive cells following receptor binding and receptor dimerization. The dimerization of VEGF receptors causes receptor autophosphorylation, which in turn activates MAP kinase (MAPK) intracellular pathways. For example, VEGF, after forming a homodimer, binds to two KDR molecules, for example, at immunoglobulin-like domains Ig2 and Ig3 of KDR. This results in KDR dimerization and activation.
It is widely accepted that tumor growth beyond a few cubic millimeters cannot occur without inducing a new vascular supply. Thus, inhibiting the development of new blood vessels is a potential approach to cancer therapy that has attracted interest in recent years. In view of the role of VEGF in vascular endothelial proliferation and angiogenesis, and the role that these processes play in many different diseases and disorders, it is desirable to have a means for reducing or inhibiting one or more of the biological activities of VEGF. Currently, blockade of the VEGF pathway is achieved by many different means, including blocking antibodies targeted against VEGF or its receptors (Dvorak, HF, J. Clin. Oncology, 20: 4368-4380 (2002)), soluble decoy receptors that prevent VEGF from binding to its normal receptors (Kim et al., PNAS U.S.A., 99: 11399-11404 (2002)), and small molecule inhibitors of tyrosine kinase activity of the VEGF receptors.
The present inventors have identified novel compositions and methods for inhibiting VEGF signaling. Thus, the present invention provides a means for reducing or inhibiting endogenous VEGF activity and, in turn, reducing or inhibiting endothelial cell proliferation and angiogenesis.